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The activity and expression of DNA repair/photoreactivation with the enzyme photolyase in the sea urchin Sterechinus neumayeri, McMurdo Sound

Entry ID:
K068_2003_2004_NZ_1

Summary

Abstract:
An important aspect of understanding the impact of increased UVR is assessing the ability of Antarctic species to repair DNA damaged through the process of photoreactiviation or photorepair. To repair DNA damage organisms have evolved the enzyme photolyase, a light dependent enzyme that requires UVR and visible irradiance to be catalytic. To quantify the importance of photorepair rates in ... Antarctic invertebrate larvae and the importance in mitigating effects of UVR four experiments were conducted. 1. Photorepair was examined in embryos and larvae of the sea urchin Sterechinus neumayeri. Photoreactivation at the whole organism level (eggs) and at the molecular level in terms of the production and disappearance of cyclobutance pyrimidine dimers (CPDs) in DNA in eggs, blastula and 4 armed larvae was analysed. 2. Freshly fertilised S. neumayeri eggs were exposed to either 0 (control), 1 or 2 hours of UVR and the time to when 50% of the eggs have undergone first cell division recorded. Cleavage delay as a result of UV exposure can then be expressed as a percentage. Following this, a further experiment was carried out where UVR induced cleavage delay was measured in eggs where photorepair was stimulated (exposed to visible light) or inhibited (kept in the dark). 3. The production of CPDs over time was examined in the eggs, blastula and 4 armed larvae of S. neumayeri which were exposed to UVR for a total of 5 hours. Samples were taken at time 0 and at 1 hourly intervals. Embryos were fixed in buffered DMSO and later the DNA extracted and CPDs quantified. 4. To obtain a measure of the rate of photoreactivation, the disappearance of CPDs over time in S. neumayeri embryos (fertilised eggs, blastula and 4 armed larvae) that were either able to photorepair or were photorepair inhibited was examined. Embryos were exposed to UVB then allowed to photorepair by placement under a full spectrum light or total darkness (photorepair inhibited). Samples were taken at 0, 1, 2, 6, 12 and 24 hours post UV exposure. Embryos were fixed in buffered DMSO and later the DNA extracted and CPDs quantified. Identical experiments were carried out on temperate New Zealand echinoderm species and other Antarctic species to obtain a comparative rate of photoreactivation; Perknaster, Parbolarsia and Acodontaster (all freshly fertilized larvae).